This work introduces and explores a novel model which incorporates Fetal Thymus Organ Culture (FTOC) from non-obese Diabetic (NOD) mice to replicate thymic development and function of diabetogenic T cells in vitro. NOD FTOC is shown to posses a predictable diabetogenic activity measured in vitro, and a protective, regulatory activity when adoptively transferred to genetically IDDM-predisposed NOD mice. This in vitro IDDM (ivIDDM) activity is unique to NOD FTOC, and can be abbrogated by co-culture of developing NOD FTOC with FT from immunologically incompetent C.B-17 scid/scid mice. Additionally early exposure of NOD T cell precursors to islet antigens by co-culture with NOD Fetal Pancreas can negatively select for diabetogenic T cells or activate immuno-regulatory cells that can suppress diabetogenic T cell activity. The addition of blocking F(ab' )2 fragments of anti-CD3epsilon monoclonal antibody to NOD FTOC/FP co-cultures prevented insulin reduction, implicating a role for TcR-mediated recognition in this "in vitro IDDM" model. Transfer of unprimed syngeneic FTOC cells to pre-diabetic NOD mice prevents the onset of IDDM while transfer of islet-cell primed FTOC/FP cells slightly increased disease incidence. Spontaneous proliferation to peptides of Glutamate Decarboxylase (GAD) was not detected in NOD FTOC in contrast to reports of such responses in pre-diabetic NOD mice. A marked response to GAD peptides is induced by priming NOD FTOC, and increases ivIDDM. Proliferation is significantly diminished by tolergenic early treatment of FTOC, as is ivIDDM activity. Offspring of GAD peptide-treated NOD mice have a lower incidence of IDDM indicating potential beneficial tolerance to islet antigens by in utero exposure. Injection of identical GAD peptides to pre-diabetic NOD mice enhances the incidence of IDDM demonstrating the deleterious effects of inappropriate in vivo administration of autoantigenic peptides. NOD FTOC is shown to readily integrate and express retrovirus-delivered class II MHC I-E(α)d as measured by PCR and flow cytometry, respectively. The contribution to understanding the antigenic, genetic and regulatory basis of IDDM in NOD mice and humans is discussed.

This work introduces and explores a novel model which incorporates Fetal Thymus Organ Culture (FTOC) from non-obese Diabetic (NOD) mice to replicate thymic development and function of diabetogenic T cells in vitro. NOD FTOC is shown to posses a predictable diabetogenic activity measured in vitro, and a protective, regulatory activity when adoptively transferred to genetically IDDM-predisposed NOD mice. This in vitro IDDM (ivIDDM) activity is unique to NOD FTOC, and can be abbrogated by co-culture of developing NOD FTOC with FT from immunologically incompetent C.B-17 scid/scid mice. Additionally early exposure of NOD T cell precursors to islet antigens by co-culture with NOD Fetal Pancreas can negatively select for diabetogenic T cells or activate immuno-regulatory cells that can suppress diabetogenic T cell activity. The addition of blocking F(ab' )2 fragments of anti-CD3epsilon monoclonal antibody to NOD FTOC/FP co-cultures prevented insulin reduction, implicating a role for TcR-mediated recognition in this "in vitro IDDM" model. Transfer of unprimed syngeneic FTOC cells to pre-diabetic NOD mice prevents the onset of IDDM while transfer of islet-cell primed FTOC/FP cells slightly increased disease incidence. Spontaneous proliferation to peptides of Glutamate Decarboxylase (GAD) was not detected in NOD FTOC in contrast to reports of such responses in pre-diabetic NOD mice. A marked response to GAD peptides is induced by priming NOD FTOC, and increases ivIDDM. Proliferation is significantly diminished by tolergenic early treatment of FTOC, as is ivIDDM activity. Offspring of GAD peptide-treated NOD mice have a lower incidence of IDDM indicating potential beneficial tolerance to islet antigens by in utero exposure. Injection of identical GAD peptides to pre-diabetic NOD mice enhances the incidence of IDDM demonstrating the deleterious effects of inappropriate in vivo administration of autoantigenic peptides. NOD FTOC is shown to readily integrate and express retrovirus-delivered class II MHC I-E(α)d as measured by PCR and flow cytometry, respectively. The contribution to understanding the antigenic, genetic and regulatory basis of IDDM in NOD mice and humans is discussed.

en_US

dc.type

text

en_US

dc.type

Dissertation-Reproduction (electronic)

en_US

dc.subject

Health Sciences, Immunology.

en_US

thesis.degree.name

Ph.D.

en_US

thesis.degree.level

doctoral

en_US

thesis.degree.discipline

Graduate College

en_US

thesis.degree.discipline

Microbiology and Immunology

en_US

thesis.degree.grantor

University of Arizona

en_US

dc.contributor.advisor

DeLuca, Dominick

en_US

dc.identifier.proquest

9806825

en_US

dc.identifier.bibrecord

.b37555819

en_US

All Items in UA Campus Repository are protected by copyright, with all rights reserved, unless otherwise indicated.